minocycline and Diabetic-Retinopathy

Inflammation contributes significantly to the pathogenesis of diabetic macular edema (DME). In particular, retinal microglia demonstrate increased activation and aggregation in areas of DME. Study authors investigated the safety and potential efficacy of oral minocycline, a drug capable of inhibiting microglial activation, in the treatment of DME.. A single-center, prospective, open-label phase I/II clinical trial enrolled five participants with fovea-involving DME who received oral minocycline 100 mg twice daily for 6 months. Main outcome measurements included best-corrected visual acuity (BCVA), central retinal subfield thickness (CST), and central macular volume using spectral domain optical coherence tomography (SD-OCT) and late leakage on fluorescein angiography (FA).. Findings indicated that the study drug was well tolerated and not associated with significant safety issues. In study eyes, mean BCVA improved continuously from baseline at 1, 2, 4, and 6 months by +1.0, +4.0, +4.0, and +5.8 letters, respectively, while mean retinal thickness (CST) on OCT decreased by -2.9%, -5.7%, -13.9, and -8.1% for the same time points. At month 6, mean area of late leakage on FA decreased by -34.4% in study eyes. Mean changes in contralateral fellow eyes also demonstrated similar trends. Improvements in outcome measures were not correlated with concurrent changes in systemic factors.. In this pilot proof-of-concept study of DME, minocycline as primary treatment was associated with improved visual function, central macular edema, and vascular leakage, comparing favorably with historical controls from previous studies. Microglial inhibition with oral minocycline may be a promising therapeutic strategy targeting the inflammatory etiology of DME. (ClinicalTrials.gov number, NCT01120899.).

Topics: Administration, Oral; Aged; Anti-Bacterial Agents; Diabetes Mellitus, Type 2; Diabetic Retinopathy; Female; Fluorescein Angiography; Glycated Hemoglobin; Humans; Macular Edema; Male; Middle Aged; Minocycline; Prospective Studies; Retina; Tomography, Optical Coherence; Treatment Outcome; Visual Acuity

This study evaluated the safe dosage of minocycline hydrochloride (Mino) eye drops and investigated the potential for the prevention or reduction of retinal damage in a diabetic rat model.. Mino eye drops at 1 mg/ml were safe when used in SD rats. Mino eye drops can protect the retina from the development or progression of diabetic retinopathy.

Topics: Animals; Diabetic Retinopathy; Electroretinography; Humans; Minocycline; Ophthalmic Solutions; Rats; Rats, Sprague-Dawley; Retina

In this study we quantified the alterations of retinal histone post-translational modifications (PTMs) in diabetic rats using a liquid chromatography - tandem mass spectrometry (LC-MS/MS) approach. Some diabetic rats were subsequently treated with minocycline, a tetracycline antibiotic, which has been shown to inhibit the diabetes-induced chronic inflammation in the retinas of rodents. We quantified 266 differentially modified histone peptides, including 48 out of 83 methylation marks with significantly different abundancein retinas of diabetic rats as compared to non-diabetic controls. About 67% of these marks had their relative abundance restored to non-diabetic levels after minocycline treatment. Mono- and di-methylation states of histone H4 lysine 20 (H4K20me1/me2), markers related to DNA damage response, were found to be up-regulated in the retinas of diabetic rats and restored to control levels upon minocycline treatment. DNA damage response biomarkers showed the same pattern once quantified by western blotting. Collectively, this study indicates that alteration of some histone methylation levels is associated with the development of diabetic retinopathy in rodents, and the beneficial effect of minocycline on the retinas of diabetic rodents is partially through its ability to normalize the altered histone methylation levels.

Topics: Animals; Anti-Bacterial Agents; Cell Culture Techniques; Diabetic Retinopathy; DNA Damage; Histone Code; Histones; Methylation; Minocycline; Protein Processing, Post-Translational; Rats; Rats, Sprague-Dawley; Retina

Chronic low-grade inflammation occurs in diabetic retinopathy (DR), but the underlying mechanism(s) remains (remain) unclear. NLRP3 inflammasome activation is involved in several other inflammatory diseases. Thus, we investigated the role of the NLRP3 inflammasome signaling pathway in the pathogenesis of DR.. Diabetes was induced in rats by streptozotocin treatment for 8 weeks. They were treated with NLRP3 shRNA or minocycline during the last 4 weeks. High glucose-exposed human retinal microvascular endothelial cells (HRMECs) were co-incubated with antioxidants or subjected to TXNIP or NLRP3 shRNA interference.. In high glucose-exposed HRMECs and retinas of diabetic rats, mRNA and protein expression of NLRP3, ASC, and proinflammatory cytokines were induced significantly by hyperglycemia. Upregulated interleukin (IL)-1β maturation, IL-18 secretion, and caspase-1 cleavage were also observed with increased cell apoptosis and retinal vascular permeability, compared with the control group. NLRP3 silencing blocked these effects in the rat model and HRMECs, confirming that inflammasome activation contributed to inflammation in DR. TXNIP expression was increased by reactive oxygen species (ROS) overproduction in animal and cell models, whereas antioxidant addition or TXNIP silencing blocked IL-1β and IL-18 secretion in high glucose-exposed HRMECs, indicating that the ROS-TXNIP pathway mediates NLRP3 inflammasome activation. Minocycline significantly downregulated ROS generation and reduced TXNIP expression, subsequently inhibited NLRP3 activation, and further decreased inflammatory factors, which were associated with a decrease in retinal vascular permeability and cell apoptosis.. Together, our data suggest that the TXNIP/NLRP3 pathway is a potential therapeutic target for the treatment of DR, and the use of minocycline specifically for such therapy may be a new avenue of investigation in inflammatory disease.

Topics: Animals; Anti-Inflammatory Agents; Apoptosis; Capillary Permeability; Carrier Proteins; Caspase 1; Caspase 3; Cell Cycle Proteins; Cells, Cultured; Diabetes Mellitus, Experimental; Diabetic Retinopathy; Endothelial Cells; Glucose; Humans; Inflammasomes; Inflammation; Interleukin-18; Interleukin-1beta; Male; Minocycline; NLR Family, Pyrin Domain-Containing 3 Protein; Rats, Sprague-Dawley; Reactive Oxygen Species; RNA, Small Interfering

The present study aimed to investigate the mechanism underlying the effects of minocycline on diabetic retinopathy‑associated cellular apoptosis. A total of 40 Sprague Dawley (SD) rats were used as a diabetic retinopathy model following injection with streptozotocin. Among the 34 rats in which the diabetes model was successfully established, 24 rats were divided into two experimental groups: I and II (T1 and T2, respectively), and orally administered with various doses of minocycline. The remaining 10 rats served as the diabetic retinopathy control group. An additional group of 10 healthy SD rats with comparable weight served as normal controls. The rats in T1 and T2 groups were treated daily for eight consecutive weeks with minocycline at a dose of 2.5 mg/kg and 5 mg/kg, respectively. The mRNA expression levels of poly (ADP‑ribose) polymerase‑1 (PARP‑1) were subsequently measured by reverse transcription‑quantitative polymerase chain reaction, and the protein expression levels of poly‑ADP‑ribose were measured by western blot analysis and immunohistochemistry. Retinal morphology was observed following hematoxylin and eosin staining, and retinal cell apoptosis was measured by terminal deoxynucleotidyl transferase dUTP nick end labeling and caspase‑3 activity assays. The amplitudes of the electroretinogram (ERG) b‑wave and oscillary potentials (OPs) were measured using visual electrophysiology, and compared among the four groups. The results of the present study demonstrated that in the diabetic rats, retinal PARP‑1 gene expression was markedly upregulated, the number of apoptotic cells and the activity levels of caspase‑3 were increased, and the amplitude of the ERG b‑wave and the OPs were markedly lower as compared with the normal rats. Following treatment with minocycline, the abnormal expression of PARP‑1 in the retina was inhibited, and cellular apoptosis was decreased. In conclusion, the results of the present study suggest that PARP‑1 is involved in the development of diabetic retinopathy, and minocycline is able to inhibit PARP‑1 expression and decrease cellular apoptosis, suggesting that minocycline may prove to be a promising drug for the treatment of diabetic retinopathy.

Topics: Animals; Apoptosis; Caspase 3; Diabetes Mellitus, Experimental; Diabetic Retinopathy; Disease Models, Animal; Gene Expression Regulation; In Situ Nick-End Labeling; Male; Minocycline; Poly (ADP-Ribose) Polymerase-1; Poly(ADP-ribose) Polymerases; Rats; Rats, Sprague-Dawley; Retina; Streptozocin

Histone acetylation was significantly increased in retinas from diabetic rats, and this acetylation was inhibited in diabetics treated with minocycline, a drug known to inhibit early diabetic retinopathy in animals. Histone acetylation and expression of inflammatory proteins that have been implicated in the pathogenesis of diabetic retinopathy were increased likewise in cultured retinal Müller glia grown in a diabetes-like concentration of glucose. Both the acetylation and induction of the inflammatory proteins in elevated glucose levels were significantly inhibited by inhibitors of histone acetyltransferase (garcinol and antisense against the histone acetylase, p300) or activators of histone deacetylase (theophylline and resveratrol) and were increased by the histone deacetylase inhibitor, suberolylanilide hydroxamic acid. We conclude that hyperglycemia causes acetylation of retinal histones (and probably other proteins) and that the acetylation contributes to the hyperglycemia-induced up-regulation of proinflammatory proteins and thereby to the development of diabetic retinopathy.

Topics: Acetylation; Amino Acid Sequence; Animals; Cell Line; Diabetes Mellitus, Experimental; Diabetic Retinopathy; Histone Acetyltransferases; Histone Deacetylase Inhibitors; Histone Deacetylases; Histones; Inflammation Mediators; Intercellular Adhesion Molecule-1; Male; Minocycline; Molecular Sequence Data; Nitric Oxide Synthase Type II; Peptide Fragments; Protein Processing, Post-Translational; Rats; Rats, Inbred Lew; Retina; Tandem Mass Spectrometry; Terpenes; Vascular Endothelial Growth Factor A

Inflammation plays important roles in the development of diabetic retinopathy (DR). How Müller cells contribute to DR-related inflammation remains unclear. We hypothesized that under diabetic conditions, elevated histone acetylations in Müller cells contribute to the inflammatory response. In this study, significantly increased histone acetylations, elevated histone acetyltranferases levels, and decreased histone deacetylases levels were found in the retinas of diabetic rats. Elevated AcH3K9 and AcH3K18 were partially co-stained with Müller cells on retinal sections by immunofluorescence staining. Consistently, high-glucose (HG) treated rMC-1 cells, a Müller cell line, also showed upregulation of acetylated histones, accompanied with the overexpression of GFAP, p-STAT3, and NFκB-p65, and two inflammatory genes, TNFα and MCP-1. Meanwhile, sodium butyrate (NaB)-induced upregulation of acetylated histones is also accompanied with transcription of inflammatory genes. Minocycline, a drug with beneficial effects on DR, was found to downregulate HG-induced Müller cell activation, inflammation, and acetylated H3K18 bound to the promoters of GFAP and inflammatory genes by chromatin immunoprecipitation assay. Furthermore, the effects of minocycline on HG-induced elevation in histone acetylations were also demonstrated in isolated primary rat Müller cells. These findings suggest the elevation of histone acetylations in Müller cells plays important regulating roles in the inflammatory response during diabetic conditions. Inhibition of histone acetylation by minocycline is a novel function that may contribute to its beneficial effects on DR.

Topics: Acetylation; Animals; Cells, Cultured; Diabetes Mellitus, Experimental; Diabetic Retinopathy; Histones; Inflammation; Inflammation Mediators; Male; Minocycline; Rats; Rats, Sprague-Dawley; Retina

This study examined aquaporin 4 (AQP4) and Kir4.1 (a potassium channel subunit) in normal and diabetic adult Sprague-Dawley rats, and determined the effect of minocycline treatment. Retinal expression of the AQP4 and Kir4.1 genes was examined using double immuno fluorescence, Western blot analysis, and real-time reverse transcription-polymerase chain reaction. Retinal levels of vascular endothelial growth factor (VEGF), ionized calcium-binding adaptor molecule (Iba)-1 and interleukin (IL)-1β were also ascertained. The blood-retinal barrier (BRB) and retinal oedema were assessed using rhodamine isothiocyanate. AQP4, VEGF, Iba-1, and IL-1β mRNA and protein levels increased, and Kir4.1 mRNA and protein levels decreased, in diabetic rat retinas. Both BRB disruption and retinal oedema were also observed in these retinas. In diabetic rats, minocycline treatment decreased AQP4, VEGF, Iba-1 and IL-1β levels and retinal oedema, and increased Kir4.1 levels. These findings suggest that minocycline might be beneficial for retinal fluid clearance and reduction of retinal oedema in diabetic retinopathy.

Topics: Animals; Aquaporin 4; Blotting, Western; Diabetic Retinopathy; Fluorescent Antibody Technique; Gene Expression Regulation; Glial Fibrillary Acidic Protein; Male; Minocycline; Potassium Channels, Inwardly Rectifying; Rats; Rats, Sprague-Dawley; Retina; Reverse Transcriptase Polymerase Chain Reaction; Rhodamines; RNA, Messenger

The proinflammatory cytokine, interleukin (IL)-1beta, is known to induce vascular dysfunction and cell death. We investigated the role of IL-1beta and caspase-1 (the enzyme that produces it) in diabetes-induced degeneration of retinal capillaries. Caspase-1 activity is increased in retinas of diabetic and galactosemic mice and diabetic patients. First, we investigated the effect of agents known to inhibit caspase-1 (minocycline and tetracycline) on IL-1beta production and retinal capillary degeneration in diabetic and galactose-fed mice. Second, we examined the effect of genetic deletion of the IL-1beta receptor on diabetes-induced caspase activities and retinal capillary degeneration. Diabetic and galactose-fed mice were injected intraperitoneally with minocycline or tetracycline (5 mg/kg). At 2 months of diabetes, minocycline inhibited hyperglycemia-induced caspase-1 activity and IL-1beta production in the retina. Long-term administration of minocycline prevented retinal capillary degeneration in diabetic (6 months) and galactose-fed (13 months) mice. Tetracycline inhibited hyperglycemia-induced caspase-1 activity in vitro but not in vivo. Mice deficient in the IL-1beta receptor were protected from diabetes-induced caspase activation and retinal pathology at 7 months of diabetes. These results indicate that the caspase-1/IL-1beta signaling pathway plays an important role in diabetes-induced retinal pathology, and its inhibition might represent a new strategy to inhibit capillary degeneration in diabetic retinopathy.

Topics: Animals; Capillaries; Caspase 1; Caspase 3; Caspase Inhibitors; Diabetes Mellitus, Experimental; Diabetic Retinopathy; Galactose; Galactosemias; Glucose; Interleukin-1beta; Male; Mice; Mice, Inbred C57BL; Minocycline; Models, Animal; Retinal Degeneration; Retinal Vessels; Signal Transduction; Tetracycline

Diabetes leads to vascular leakage, glial dysfunction, and neuronal apoptosis within the retina. The goal of the studies reported here was to determine the role that retinal microglial cells play in diabetic retinopathy and assess whether minocycline can decrease microglial activation and alleviate retinal complications. Immunohistochemical analyses showed that retinal microglia are activated early in diabetes. Furthermore, mRNAs for interleukin-1beta and tumor necrosis factor-alpha, proinflammatory mediators known to be released from microglia, are also increased in the retina early in the course of diabetes. Using an in vitro bioassay, we demonstrated that cytokine-activated microglia release cytotoxins that kill retinal neurons. Furthermore, we showed that neuronal apoptosis is increased in the diabetic retina, as measured by caspase-3 activity. Minocycline represses diabetes-induced inflammatory cytokine production, reduces the release of cytotoxins from activated microglia, and significantly reduces measurable caspase-3 activity within the retina. These results indicate that inhibiting microglial activity may be an important strategy in the treatment of diabetic retinopathy and that drugs such as minocycline hold promise in delaying or preventing the loss of vision associated with this disease.

Topics: Animals; Caspase 3; Caspases; Cytokines; Diabetes Mellitus, Experimental; Diabetic Retinopathy; Disease Models, Animal; Inflammation; Microglia; Minocycline; Rats; Rats, Sprague-Dawley

Retinal neurodegenerative disease involves an inflammatory response in the retina characterized by an increase in inflammatory cytokines and activation of microglia. The degree of microglia activation may influence the extent of retinal injury following an inflammatory stimulus. Cytokines released by activated microglia regulate the influx of inflammatory cells to the damaged area. Thus, a therapeutic strategy to reduce cytokine expression in microglia would be neuroprotective. Minocycline, a semisynthetic tetracycline derivative, is known to protect rodent brain from ischemia and to inhibit microglial activation. In this study, we activated retinal microglia in culture with lipopolysaccharide (LPS) and attempted to determine whether minocycline could reduce the production of cytokines from activated microglia at both gene and protein levels. Changes in inflammatory cytokines, TNF-alpha and IL-1beta, were measured by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA) in the presence or absence of LPS. We also measured the levels of nitric oxide (NO) by the nitrate reductase method under similar conditions. LPS treatment induced a significant upregulation of the mRNA and release of TNF-alpha, IL-1beta, and NO from retinal microglia. Minocycline inhibited these releases. Thus, minocycline might exert its antiinflammatory effect on microglia by inhibiting the expression and release of TNF-alpha, IL-1beta, and NO.

Topics: Animals; Animals, Newborn; Anti-Bacterial Agents; Anti-Inflammatory Agents; Cells, Cultured; Cytokines; Diabetic Retinopathy; Encephalitis; Gliosis; Interleukin-1; Lipopolysaccharides; Microglia; Minocycline; Neuroprotective Agents; Nitric Oxide; Rats; Rats, Sprague-Dawley; Retina; Tumor Necrosis Factor-alpha; Up-Regulation